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Imaging

Identify and localize immune cells in the microenvironment of your tissues samples


Are you interested in quantitating immune cells that infiltrate the human tissues you study, understanding where they are located, or determining which other cells they are near in the tissue architecture? HIMSR operates a state-of-the-art imaging facility for many of your imaging needs. We are equipped with Akoya Biosciences’ Vectra 3.0 and Vectra Polaris, automated microscopes that images brightfield or immunofluorescence slides. We also offer IONpath’s MIBI (Multiplexed Ion Bean Imaging) technology. We have pre-developed antibody panels for Vectra and MIBI or can help develop new antibodies to help you analyze your tissues. 

These technologies are optimal for analysis of formalin-fixed, paraffin-embedded tissues or tissue microarrays (not frozen tissues). The antibodies are resolved with Akoya Opal technology, which allows use of the best primary antibodies for immunohistochemistry, regardless of species with no cross-reactivity. Log into iLabs for fees; contact Kim Jordan for a quote.


Vectra® 3
With this high-throughput instrument, we can distinguish nuclear DNA plus up to 6 different cellular markers in regions of interest. We have over 100 panels with more than 80 different human antibodies and over 10 panels with 20 mouse markers to choose from. For every antibody panel, we include stained tonsil-tissue controls, scan slides with up to a 20x objective, and can provide analysis depending on your needs.

Vectra Polaris​
With this high-throughput instrument, we can distinguish 7 colors in a full slide scan or 9 colors in regions of interest using up to a 40x objective. We are transferring many of the panels used for analysis with the Vectra 3.0 to the Polaris.


Opal technolo​gy
Opal technology includes 6 fluorophors (for Vectra 3.0) or 8 fluorophors (for Vectra Polaris) and enhanced DAPI to stain DNA which are read at different wavelengths. Briefly, we apply a primary detecting antibody, a secondary antibody, the Opal reagent, microwave to remove the antibodies but not the Opal reagent, repeat.
inForm software
inForm is a patented image analysis software package that uses trainable algorithms in a user-friendly interface to accurately visualize and quantify biomarkers in tissue sections. Capability includes phenotyping, co-localization, co-expression, tissue region detection (e.g. tumor versus stroma), and traditional scoring functions.  This software is available for use on a dedicated analysis station within the HIMSR (sign up for time on the iL​ab calendar) or, for larger projects, check out a copy of inform from the HIMSR ($). We offer periodic workshops to help you get started with inForm and Phenoptr Reports and Akoya Biosciences’ also have great new videos on how to use these softwares [inForm and Phenoptr Reports].



Leica Aut​ostainer
Using the Leica Autostainer, we have developed standardized antibody panels for the analysis of tissue microenvironments in human FFPE tissue for detection by the Vectra instrument.  The Leica Autostainer can perform the entire 7-color protocol on 30 slides in one overnight batch; 9-color protocols take 1.5 days.  We can also run smaller batches of slides or multiple different panels at one time. If you wish to perform the staining in your own laboratory, we are happy to provide the antibody clone names and staining protocols.
 
Custom IF antibody panel
If your antibody panel of choice is not yet available, we can work with you to develop a custom human antibody panel either in your lab or HIMSR. In the spirit of fostering collaborative research at CU|AMC, antibody panels developed in HIMSR will be shared with the campus as future standard panels.

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MIBI
The Multiplexed Ion Beam Imaging technology is open for business at the HIMSR! Using rare-earth metals and mass-spec for visualization, the MIBI can simultaneously distinguish over 40 markers (depends on number of metals) with high sensitivity (up to 5 log dynamic range depending on antibody) and high resolution (500 nm). Get the most from your FFPE tissue microarrays using this unique discovery tool! We currently have a 27-marker panel, but are happy to discuss customization. We use MIBItracker software to visualize the tissues.
 
See these publications and a recent tumor immunology presentation from HIMSR for details. If you’re interested in analyzing your samples with the MIBI technology contact Kim Jordan​ for additional experimental information and budget information.